Role of protein kinase C and NF-kappa B in proteolysis-inducing factor-induced proteasome expression in C2C12 myotubes

Proteolysis-inducing factor (PIF) is a sulphated glycoprotein produced by cachexia-inducing tumours, which initiates muscle protein degradation through an increased expression of the ubiquitin–proteasome proteolytic pathway. The role of kinase C (PKC) in PIF-induced proteasome expression has been studied in murine myotubes as a surrogate model of skeletal muscle. Proteasome expression induced by PIF was attenuated by 4alpha-phorbol 12-myristate 13-acetate (100 nM) and by the PKC inhibitors Ro31-8220 (10 muM), staurosporine (300 nM), calphostin C (300 nM) and Gö 6976 (200 muM). Proteolysis-inducing factor-induced activation of PKCalpha, with translocation from the cytosol to the membrane at the same concentration as that inducing proteasome expression, and this effect was attenuated by calphostin C. Myotubes transfected with a constitutively active PKCalpha (pCO2) showed increased expression of proteasome activity, and a longer time course, compared with their wild-type counterparts. In contrast, myotubes transfected with a dominant-negative PKCalpha (pKS1), which showed no activation of PKCalpha in response to PIF, exhibited no increase in proteasome activity at any time point. Proteolysis-inducing factor-induced proteasome expression has been suggested to involve the transcription factor nuclear factor-kappaB (NF-kappaB), which may be activated through PKC. Proteolysis-inducing factor induced a decrease in cytosolic I-kappaBalpha and an increase in nuclear binding of NF-kappaB in pCO2, but not in pKS1, and the effect in wild-type cells was attenuated by calphostin C, confirming that it was mediated through PKC. This suggests that PKC may be involved in the phosphorylation and degradation of I-kappaBalpha, induced by PIF, necessary for the release of NF-kappaB from its inactive cytosolic complex.

Expression of VEGF-C and activation of its receptors VEGFR-2 and VEGFR-3 in trophoblast

Placental villous development requires the co-ordinated action of angiogenic factors on both endothelial and trophoblast cells. Like vascular endothelial growth factor (VEGF), VEGF-C increases vascular permeability, stimulates endothelial cell proliferation and migration. In the present study, we investigated the expression of VEGF-C and its receptors VEGFR-3 and VEGFR-2 in normal and intrauterine growth-restricted (IUGR) placenta. Immunolocalisation studies showed that like VEGF and VEGFR-1, VEGF-C, VEGFR-3 and VEGFR-2 co-localised to the syncytiotrophoblast, to cells in the maternal decidua, as well as to the endothelium of the large placental blood vessels. Western blot analysis demonstrated a significant decrease in placental VEGF-C and VEGFR-3 protein expression in severe IUGR as compared to gestationally-matched third trimester pregnancies. Conditioned medium from VEGF-C producing pancreatic carcinoma (Suit-2) and endometrial epithelial (Hec-1B) cell lines caused an increased association of the phosphorylated extracellular signal regulated kinase (ERK) in VEGFR-3 immunoprecipitates from spontaneously transformed first trimester trophoblast cells. VEGF121 caused dose-dependant phosphorylation of VEGFR-2 in trophoblast cells as well as stimulating DNA synthesis. In addition, premixing VEGF165 with heparin sulphate proteoglycan potentiated trophoblast proliferation and the association of phospho-ERK with the VEGFR-2 receptor. VEGF165-mediated DNA synthesis was inhibited by anti-VEGFR-2 neutralising antibody. The results demonstrate functional VEGFR-2 and VEGFR-3 receptors on trophoblast and suggest that the decreased expression of VEGF-C and VEGFR-3 may contribute to the abnormal villous development observed in IUGR placenta.

Crystal structure of [{Cu(H2O)(en)2}{Cu(en)2}Re6Te8(CN)6]·3H2O

The reaction of Cs4[Re6Te8(CN)6]·2H2O with Cu(en)2Cl2 in water affords crystals of a cluster complex [{Cu(H2O)(en) 2}{Cu(en)2}Re6Te8(CN)6]·3H2O. The structure of the compound is determined by single crystal X-ray diffraction (a = 10.8082(4) Å, b = 16.5404(6) Å, c = 24.6480(7) Å, β = 92.696(1)°, V = 4401.5(3) Å3, Z = 4, space group P21/n, R 1 = 0.0331, wR 2 (all data) = 0.0652). In the complex, cluster [Re6Te8(CN)6]4- anions are linked by Cu2+ cations into zigzag chains through cyanide bridges. The coordination environment of the copper cations is complemented by ethylenediamine molecules. Each of the cluster anions is additionally coordinated by a terminal fragment {Cu(H2O)(en)2}. © 2014 Pleiades Publishing, Ltd.